Am J Physiol Cell Physiol AJP: Heart and Circulatory Physiology
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Am J Physiol Cell Physiol 273: C1889-C1899, 1997;
0363-6143/97 $5.00
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Vol. 273, Issue 6, C1889-C1899, December 1997

Regulation of a cloned epithelial Na+ channel by its beta - and gamma -subunits

Mouhamed S. Awayda1, Albert Tousson2, and Dale J. Benos3

1 Department of Medicine and Department of Physiology, Tulane University School of Medicine, New Orleans, Louisiana 70112; and 2 Department of Cell Biology and 3 Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35223

Using the Xenopus oocyte expression system, we examined the mechanisms by which the beta - and gamma -subunits of an epithelial Na+ channel (ENaC) regulate alpha -subunit channel activity and the mechanisms by which beta -subunit truncations cause ENaC activation. Expression of alpha -ENaC alone produced small amiloride-sensitive currents (-43 ± 10 nA, n = 7). These currents increased >30-fold with the coexpression of beta - and gamma -ENaC to -1,476 ± 254 nA (n = 20). This increase was accompanied by a 3.1- and 2.7-fold increase of membrane fluorescence intensity in the animal and vegetal poles of the oocyte, respectively, with use of an antibody directed against the alpha -subunit of ENaC. Truncation of the last 75 amino acids of the beta -subunit COOH terminus, as found in the original pedigree of individuals with Liddle's syndrome, caused a 4.4-fold (n = 17) increase of the amiloride-sensitive currents compared with wild-type alpha beta gamma -ENaC. This was accompanied by a 35% increase of animal pole membrane fluorescence intensity. Injection of a 30-amino acid peptide with sequence identity to the COOH terminus of the human beta -ENaC significantly reduced the amiloride-sensitive currents by 40-50%. These observations suggest a tonic inhibitory role on the channel's open probability (Po) by the COOH terminus of beta -ENaC. We conclude that the changes of current observed with coexpression of the beta - and gamma -subunits or those observed with beta -subunit truncation are likely the result of changes of channel density in combination with large changes of Po.

oocyte expression; immunofluorescence; Liddle's syndrome; channel activation


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